Current status and perspectives of genome editing technology for microalgae

Jeon, Seungjib; Lim, Jong-Min; Lee, Hyung-Gwan; Shin, Sung-Eun; Kang, Nam Kyu; Park, Youn-Il; Oh, Hee-Mock; Jeong, Won-Joong; Jeong, Byeong-ryool; Chang, Yong Keun

doi:10.1186/s13068-017-0957-z

Biotechnology for Biofuels and Bioproducts

Table 2 Genome editing in microalgae and other organisms

From: Current status and perspectives of genome editing technology for microalgae

Organism	Delivery/nuclease	Gene/marker	Comments	References
Animals
CHO-S cells (11619-012)	Lipofectamine 2000 transfection ZFN	DHFR	Identification of biallelic knockouts after a single transient ZFN treatment	[57]
SKOV3 cells ATCC	FuGENE HD transfection reagent TALEN	EGFP and DsRed	Development of TALENs in mammalian cells	[59]
Mouse cells NIH3T3	Microinjection or Lipofectamine 2000 CRISPR/Cas9	EGFP and ApoE	Improvement of bi-allelic modification by dual sgRNAs	[178]
Human cell lines MCF-7, NSCLC and A549	Lipofectamine 2000 transfection CRISPR/Cas9	Mitochondrial GTPase Mitofusin-2/GFP	Identification of role MFN2 in human cells	[179]
H9 human ES cells and other cell lines	Electroporation CRISPR/Cas9	CCR5 gene	Genome editing Cas9 RNPs	[65]
Mouse zygotes	Electroporation CRISPR/Cas9	Tyrosinase gene	Genome editing Cas9 RNPs	[180]
Zebrafish (Danio rerio) strain AB	Microinjection into embryos CRISPR/Cas9	akt2 gene	Functional role of akt2 gene in zebrafish	[181]
Plants
Nicotiana tabacum (tobacco)	Protoplast transformation TALEN	ALS gene/YFP gene	Successful targeted gene replacement with TALENs	[51]
Arabidopsis thaliana	Agrobacterium-mediated transformation CRISPR/Cas9	ADH1, TT4, RTEL1 gene/bar gene	Genome editing using CRISPR/Cas9-based nucleases and nickases	[101]
Arabidopsis thaliana	Agrobacterium-mediated transformation CRISPR/Cas9	GFP gene	Stable inheritance of Cas9/sgRNA-generated mutant genes in T2 and T3 progeny	[182]
Camelina sativa	Agrobacterium-mediated transformation CRISPR/Cas9	FAD2 gene	Induced mutations caused change in fatty acid composition	[183]
Maize Hi-line	Particle bombardment CRISPR/Cas9	LIG, ALS2, MS26 and MS45/MOPAT-DSREB	Genome editing Cas9 RNPs	[184]
Wheat embryos	Particle bombardment CRISPR/Cas9	TaGW2 gene	Genome editing Cas9 RNPs	[185]
Cyanobacteria
Synechococcus elongatus UTEX 2973	Conjugation CRISPR/Cas9	nblA	Genome editing using CRISPR/Cas9-based nucleases	[186]
Synechococcus elongatus PCC 7942	Conjugation CRISPR/Cas9	glgc/Gm^R gene	Increase of succinate Production	[187]
Synechococcus UTEX 2973	Conjugation CRISPR/Cpf1	psbA1, nblA	Genome editing using CRISPR/Cpf1-based nucleases	[188]
Synechocystis 6803 Anabaena 7120	Conjugation CRISPR/Cpf1	nblA nifH	Genome editing using CRISPR/Cpf1-based nucleases	[188]
Microalgae
C. strain CC-4350	Glass beads ZFN	COP3 gene/aphVIII gene	Targeted gene knockout induced by ZFN	[58]
P. tricornutum CCMP2561	Bombardment TALEN	UGPase/NAT gene	Increase in triacylglycerol accumulation	[91]
C. reinhardtii CC503	Electroporation CRISPR/Cas9	FKB12 gene	First application of CRISPR/Cas9 in microalgae	[111]
P. tricornutum CCMP2561	Bombardment CRISPR/Cas9	CpSRP54 gene/Shble	First application of CRISPR/Cas9 in diatoms	[189]
C. reinhardtii CC-124	Electroporation CRISPR/Cas9	MAA7, CpSRP43 and ChlM	Targeted gene knockout and knock-in via NHEJ in Chlamydomonas	[60]
C. reinhardtii CC-4349	Electroporation CRISPR/Cas9	ZEP and CpFTSY	Production of two-gene knockout mutant	[112]
C. reinhardtii CC-400	Glass beads CRISPR/dCas9	PEPC1 and RFP	CRISPRi in Chlamydomonas	[104]
N. oceanica IMET1	Electroporation CRISPR/Cas9	Nitrate reductase gene/HygR	Targeted gene knockout in Nannochloropsis	[113]
N. gaditana CCMP1894	Electroporation Cas9 Editor line	ZnCys TF BSD	Knockout and attenuation of ZnCys in Nannochloropsis	[114]

ADH1, alcohol dehydrogenase 1; akt2, AKT serine/threonine-protein kinases 2; ALS, acetolactate synthase; aphVIII, aminoglycoside 3′-phosphotransferase; ApoE, apolipoprotein E; bar, herbicide bialaphos; BSD, blasticidin S deaminase; CCR5, C–C motif chemokine receptor 5; ChlM, Mg-protoporphyrin IX S-adenosyl methionine O-methyl transferase; COP3, light-gated proton channel rhodopsin; CpFTSY, signal recognition particle receptor protein, chloroplast; CpSRP43, chloroplast signal recognition particle 43; CpSRP54, chloroplast signal recognition particle 54; DHFR, dihydrofolate reductase; DsRed, red fluorescent protein; EGFP, green fluorescent protein; FAD2, fatty acid desaturase 2; FKB12, peptidyl-prolyl cis–trans isomerase; glgc, glucose-1-phosphate adenylyl transferase; Gm^R, gentamycin-resistance gene; HygR, hygromycin resistance; LIG, liguleless1; MAA7, beta subunit of tryptophan synthase; MFN2, mitochondrial GTPase mitofusin-2; MS26 and MS45, male fertility genes; NAT, N-acetyl transferase; nblA, phycobilisome degradation protein; nifH, nitrogenase reductase; PEPC, phosphoenolpyruvate carboxylase; PsbA1, D1 protein of photosystem II; RFP, red fluorescent protein; RTEL1, regulator of telomere length 1; TaGW2, gene related to grain development; TT4, transparent testa 4; UGPase, UDP-glucose pyrophosphorylase; YFP, yellow fluorescent protein; ZEP, zeaxanthin epoxidase

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